PROJECT SUMMARY Death related to severe infection, or sepsis, is inextricably linked to the refractory failure of one or more organ systems. While advances have been made in understanding key molecular mechanisms by which specific microorganisms cause severe infection, we still have a poor understanding of how a diverse array of human pathogens elicit the rapid progression of systemic disease defined as multiple organ dysfunction syndrome (MODS). Among children with sepsis, over 90% demonstrate multi-organ dysfunction, providing a clinical imperative to understand fundamental mechanisms by which pediatric sepsis causes MODS. With rising rates of antimicrobial resistance and the refractory nature of MODS, a deeper understanding of how localized infection converts to life-threatening systemic disease is needed. We have demonstrated the role of zinc dependent metalloproteases in the pathogenesis of lethal infection caused by S. aureus and B. fragilis ? two leading etiologic agents of human sepsis. S. aureus ?-toxin utilizes host ADAM10 (A Disintegrin and Metalloprotease 10) as its cellular receptor, triggering untimely, pathologic activation of the metalloprotease. The B. fragilis toxin (BFT) is a zinc dependent metalloprotease similar to ADAM10. ADAM10 and BFT both contribute to lethal infection-associated MODS. Based on these findings, we propose that infection-associated MODS is catalyzed by aberrant activation of host ADAM10. As ADAM10 is ubiquitously expressed and involved in diverse host cellular processes, this metalloprotease is well-positioned to cause the scope of injuries observed in MODS. The primary goal of this proposal is thus to examine the hypothesis that ADAM10 is required for the development of MODS in the setting of severe pneumonia and bloodstream infection caused by key human bacterial and fungal pathogens. This proposal is based on three discoveries: 1) Cell-type specific ADAM10 knockout mice permit the identification of effects of S. aureus ?-toxin on individual cells in lethal infection, revealing that these depend on toxin-mediated activation of ADAM10 and pathologic cleavage of native ADAM10 substrates. 2) Infection-related organ dysfunction is a composite of ADAM10 action on discrete cells, integrated in the tissue over time. 3) A specific polymorphism in human ADAM10 predisposes toward the development of severe sepsis, irrespective of the inciting pathogen. Extending these observations toward the in vivo analysis of the broader role of ADAM10 in MODS as proposed will provide an opportunity to examine the innovative hypothesis that a host metalloprotease is critical for MODS. As specific inhibitors of ADAM10 have demonstrated safety in human clinical trials, knowledge gained from these studies may enable the evaluation of a novel class of therapeutics for infection-associated MODS.